Chapter 18: Fundamentals of Spectrophotometry

I. Using light in Spectroscopy

A. Properties of light

-light is described as both particles and waves

Plane polarized light..

Visible: ~ 380-780 nm UV: 160 ~ 380nm UV/Vis: 160-780nm

(these cause transitions that we care about)

Electrical component: transitions we care about (absorption emission)

Magnetic component: Absorption of rf waves (NMR)

I. Using light in Spectroscopy

A. Properties of light

I. Using light in Spectroscopy

A. Properties of light

c=νλ where c (outside vacuum)= c/nRefractive index of solution

E= hν = hc/λ = hc ν

Wavenumber (cm-1) = 1/λ

B. Absorption of Light

Electronic transitions

may result in: rotational

vibrational changes

(increase in energy of molecule)

1. What happens when molecules absorb light?

B. Absorption of Light

1. What happens when molecules absorb light?

-MO diagram of formaldehyde

-electronic transitions near 397

or 355 (UV)

-is formaldehyde green-yellow?

a. Electronic transitions

Ground

planar

Excited

pyramidal

B. Absorption of Light

1. What happens when molecules absorb light?

a. Electronic transitions

-or if IR radiation, little E

no electronic transitions, only vibrational

-vibrational and rotational transitions

usually go together

-think microwave dinner

B. Absorption of Light

1. What happens when molecules absorb light?

b. Rotational and Vibrational Transitions

-As analytical chemists these aren’t

as important to us

B. Absorption of Light

1. What happens when molecules absorb light?

c. Electronic, Rotational and Vibrational Transitions

-Electronic transitions

usually involve

simultaneous vibrational

and rotational

-most transitions radiationless

(heat)

-UV/Vis detection is

based on absorption not

emission

Ground

Excited

F= Fluorescence P=Phosphorescence

-electronic absorption bands

usually broad

-many vibrational and rotational

levels avail. at slightly different

energies

Visible absorption spectra

c. Electronic, Rotational and Vibrational Transitions

-We want to detect at UV max to get best sensitivity

D. UV/Vis instrumentation- for Absorbance!

I. Using light in Spectroscopy

Light

Source

hν

Hg, Deuterium,

Xe for UV, or W for Vis

Filter (monochrometer)

Path length Detection

(flow cell)

-Choose λ max for analytes

-Want small detector volume so little

band broadening

-Common for LC

PincidentPath length

bPtransmitted

A= log Po/P = -log T

Beer’s Law

A=єbC

II. Atomic and Molecular Spectroscopy (see Chapter 21)

II. Atomic and Molecular Spectroscopy (see Chapter 21)

A. UV/VIS (looking at molecules)

- kinetics, molar concentration determination,

need a chromophore, look at molecules

B. Atomic Absorption (AA) (looking at atoms)

Lamp flame monochromator detector

-Use lamp made of the metal detecting- shoots λ at flame

-detection of metals in a flame 2000-300K (acetylene/air)

-sample atomized in flame (the flame is the cuvet)

-one metal at a time at its max λ

-need different lamp for each metal

-measure ground excited state absorbance

II. Atomic and Molecular Spectroscopy (see Chapter 21)

B. Atomic Absorbance (AA)

To detect Fe

it’s made of Fe

LOD 0.001 to 0.020 ppm

1 to 20 ng/mL (ppb)

II. Atomic and Molecular Spectroscopy

-Absorption and emission are mirror images

-emission is at lower energy than absorption

…geometry and solvation are factors

-Inductively Coupled Plasma (ICP)

II. Atomic and Molecular Spectroscopy (see Chapter 21)

C. Atomic Emission Spectroscopy (looking at atoms)

-plasma of Ar

-no light source!

-Measure emission

Excited Ground

Advantage:

measure >1 metal/run

-detect all metals!

-60 metals simultaneously!LOD 10 ppb or less

Let’s do some calculations for UV/Vis on the board…

I. Using light in Spectroscopy Revisited (Remix)

Ex1:a) What value of absorbance corresponds to 45.0 % T?

b) If a 0.0100 M soln. exhibits 45.0 % T, what will the %T be for a

0.0200M solution of the same compound at the same wavelength?

Ex2:

a) A 3.96 x 10-4 M solution of A had an absorbance of 0.624 at 238 nm in a 1.00

cm cuvet. A blank solution containing only solvent had an absorbance of 0.029 at

the same wavelength. Find the molar absorptivity of compound A.

I. Using light in Spectroscopy Revisited (Remix)

Ex2:

b) The absorbance of an unknown amount of compound A under the same

conditions as part a was 0.375 at 238 nm. Find the concentration

of A in the unknown.